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Publication Abstract from PubMed

Type III CRISPR-Cas effectors recognize target RNAs complementary to the crRNA guide, activating diverse downstream antiviral responses. In contrast to type III-A systems, the architecture of the type III-B effector (Cmr), comprising six proteins (Cmr1-Cmr6) and a crRNA, remains incompletely defined. Moreover, although maturation of the 3' region of type III crRNA has been attributed to polynucleotide phosphorylase (PNPase), an alternative maturation pathway has been suggested but remains to be elucidated. Here we determined the cryo-EM structure of the Cmr1-lacking Archaeoglobus fulgidus Cmr (AfCmrDelta1) bound to a target analog at 3.4 A resolution. The complex forms a continuous basic channel that accommodates a crRNA-target heteroduplex. Comparative interface analysis explains why the previously reported cross-species Cmr assembly retains activity, revealing interface flexibility that enables compatible Cmr3-Cmr4 and Cmr2-Cmr5 interactions. Furthermore, we show the cooperative, site-specific processing of an intermediate crRNA that requires both AfCmrDelta1 and AfCmr1 and proceeds without divalent cations. In addition to identifying the cleavage site within the intermediate crRNA, mutational analysis of AfCmr1 reveals residues critical for the reaction. These findings suggest an alternative pathway for crRNA maturation during type III effector assembly that complements PNPase-mediated trimming of the intermediate crRNA, thereby expanding the mechanistic landscape of type III CRISPR-Cas systems.

Cryo-EM structure of Archaeoglobus fulgidus type III-B CRISPR-Cas effector and intermediate crRNA processing during effector assembly.,Ishihara K, Kitagawa S, Adachi N, Akutsu M, Senda T, Inanaga H, Numata T Biochem Biophys Res Commun. 2025 Dec 15;792:152978. doi: , 10.1016/j.bbrc.2025.152978. Epub 2025 Nov 13. PMID:41242302^[1]

From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine.